This research proposal has been stimulated by our discovery that CD38, an ectoenzyme originally thought to be exclusively localized to lymphocytes, is present in significant levels in the cell membranes of retinal Muller cells. To understand the functional role of CD38, we have developed a hypothesis, derived from our own experiments, in which we propose that external NAD, released from cells, activates CD38 to produce internalized second-messengers, such as cyclic ADP- ribose (cADPR)and/or nicotinic acid adenine dinucleotide phosphate (NAADP). Our working hypothesis is that cADPR and/or NAADP modulate the internal levels of calcium in Muller cells and that cADPR alters calcium through activation of ryanodine receptors. Activation of ryanodine receptors triggers calcium waves which can be blocked or modified by ryanodine, caffeine and thapsigargin, the latter of which depletes internal calcium stores. Calcium waves may alter the functional state of the Muller cells by releasing agents, such as ATP, which can affect neurons and glial cells. In addition, the changes in calcium may also trigger changes in the expression of proteins which alter the phenotype of the Muller cell. Such changes may contribute to the reactive nature of Muller cells in responding to mechanical or traumatic insults and in response to disease states. This proposal goes beyond issues related to the regulation of internal calcium through external NAD; we will thoroughly examine the mechanisms by which NAD is released from cells using both imaging and HPLC techniques (on-exchange HPLC) to assay NAD. We will determine the kinds of cells and the cellular mechanisms which permit cellular release of NAD, such as the possibility that gap junction hemi channels torn an NAD permissive pathway. We will also investigate whether externally produced cADPR could serve as a paracrine hormone to modify the behavior of nerve cells, using calcium imaging and external application of cADPR. Our general hypothesis, stimulated by our recent discoveries of CD38 and the action of NAD, is that CD38 serves as the epicenter for an external/internal communication pathway in which the afferent path is through NAD released from cells which, through CD38 is converted to internalized cADPR (and possibly NAADP)and that this pathway enhances the internal calcium levels of Muller cells and triggers calcium waves. The efferent pathway of this hypothesis is that cADPR, whether synthesized externally by CD38 or released from internal Muller cell storage circulates extracellularly and is internalized by neurons to serve as a feedback pathway to after calcium levels in retinal neurons and perhaps the pigment epithelial cells. Thus, this proposed pathway is one in which the magnitude and levels of calcium induced in Muller cells, provides a feedback, regulatory mechanism which engages and supports changes in neuronal calcium. If this hypothesis can be experimentally established, it will form the basis of an entirely new method by which neurons and glia support each other to stabilize retinal function.